Studies On The Dynamics And Functions Of Soybean Transmissible Floral Substances

Whether there exist transmissible flowering stimuli and inhibitors and what they are, are of importance in plant photoperiodic mechanism research. Soybean is a typical short day plant whose varieties have diverse photoperiodic responses, and some varieties are rather sensitive to photoperiods. Thus, it may be a suitable subject for the study of flowering stimulus and inhibitor. Here, by grafting experiments, flowering reversion system, quantitative real-time PCR (qRT-PCR) and in situ hybridization, accumulative effects of flowering stimuli and inhibitor were focused on, and their interactions were discussed. The expression of GmFT and the identities of floral substances were also analyzed and discussed.1. To elucidate whether there exist flowering stimuli and inhibitors, what the underlying mechanism is, and how they regulate plant development, the early-maturing soybean cultivar Heihe27and late-maturing cultivar Zigongdongdou were used as stock and scion respectively, and the composite plants were constructed in which scions and stocks had different leaf numbers in combination with defoliation, and the effect of leaf number of stock and scion on the reproductive development was observed. Under long-day (16h) conditions, Zigongdongdou scions flowered30days after being grafted onto Heihe27stocks, while its self-grafts failed to flower until the experiment finished (over120days). It suggests that under16h long-day conditions, the transmissible floral stimuli might be produced in the leaves of the early-maturing Heihe27and moved to the apical meristem of the late-maturing Zigongdongdou to induce the transition of the latter to flowering. And the effect of stocks on the flowering of scions was dependent on the number of remained leaves in stocks. It indicates that there exist an accumulative effect of the transmissible stimuli on soybean flowering. Defoliation on scions also promoted flowering and increased flower number, as suggested that some floral inhibitors should be produced in the scion leaves under long-day conditions. The number of flowers and pods on scions showed a negative correlation with the number of remained scion leaves. It indicates that floral inhibitors suppress the development of reproductive organs, and the inhibition has an accumulative effect as well. In addition, the node location where scions were defoliated also had an effect on flowering development.2. To observe the effects of stock age on scion flowering and that of scion age on its own flowering and agronomic traits, the composite plants were constructed with scions of different ages grafted on stocks of the same age or with scions of the same age grafted on stocks of different ages by interval sowing and homochronous grafting. The relationship between the production of floral stimuli in early-maturing stock and plant ages, and that between the competence to floral stimuli of late-maturing shoot apical meristem and plant ages were further analyzed. Under the long day conditions, as for the grafts of scions cv. Zigongdongdou at the same age on stocks cv Heihe27at different ages, scions exhibited different flowering states, which was related with their stock ages, that is, the older the stocks were, the more competent flowering induction was. It indicates that, as plant age increases, more mobile floral stimuli are produced. In the grafts of late scions at different ages on stock at the same age, the younger the scions were, the earlier they flowered, and the more pods were produced. Therefore, it suggests that the younger the scions are, the more sensitive they are to transmissible floral stimuli, and the sensitivity may affect some important agronomic characteristics including yield productivity.3. The spatial and temporal expression of putative florigen-encoding gene GmFT, a homologue of Flower Locus T in soybean, was analyzed in shoot apexes of late variety Zigongdongdou during vegetative growth, reproductive development and flowering reversion respectively regimed by different photoperiods. During flowering transition and reversion, GmFT transcript appeared in floral meristem and primordia of flower organ, and the expression level was related to the duration of photoperiodic treatment and reproductive development states. It indicates that GmFT’mRNA may be involved in floral meristem maintaining and organ morphogenesis, and its expression be controlled by photoperiods.4. Organ-specific, tissue-specific and diurnal expression of GmAPl, a homologue of floral meristem identity gene of API (APETALA1) in soybean, was also analyzed, in addition to bioinformatical analysis. The spatial and temporal expression of GmAP1during vegetative growth, reproductive development and flowering reversion under different photoperiods was also analyzed. GmAPl contains the domains of MADS-MEF2and K-box, typical domains for API homologues belonging to AGL8subfamily. Compared with the high expression in root, GmAPl was lowly expressed in mature leaves with a rhythmic pattern, which indicates the expression is rigorously controlled by photoperiods. In shoot apex, the expression was limited in floral meristem and primordia but with a different pattern with Arabidopsis in later organ morphogenesis.5. The expression patterns of GmFT and GmAP1were analyzed during the growth and development of varieties with gradient flowering days under LD condition, such as Heihe27, Edou4, Jilin30, Jindou19, Dian86-5, Huaxial and Zigongdongdou. SD promoted flowering of all tested varieties with GmFT and GmAPl increased at the same time. Under LD conditions, the expression of GmFT and GmAP1were negatively correlated with the flowering time of these varieties, that is, the earlier these varieties flowered, the higher the genes were expressed, which indicates that both genes must contribute to soybean maturity determination.6. To further elucidate the role of GmAPl and GmFT during flowering transition, their expression was analyzed of late-maturing scion cv. Zigongdongdou grafted on early-maturing Heihe27stock. Both GmAP1and GmFT mRNA appeared in scion, particularly in floral meristem, floral primordia and stem. The expression of GmAP1was always synchronized with the onset of scion floral meristem, which was preceded by GmFT mRNA appearance. It suggests that GmFT might act upstream of GmAPl, and its mRNA, somehow, is involved in early flowering initiation.7. Anti-GmFT antibody serum was prepared by immunizing rabbit with prokaryocyte-expressed GmFT protein. It was verified that this serum can not be used to detect GmFT protein in soybean by Western blot.Taken together, this study deepens our understanding on soybean floral substances and the expression pattern of GmFT and its related genes. In addition, molecular mechanism underlying soybean maturity diversity was also discussed. These results gave further theoretical evidences for the molecular identity of mobile floral stimuli.